A great deal of diagnostic procedures and laboratory research are carried out wherein DNA, RNA or proteins are mobilized and/or separated according to their physical and chemical properties via electrophoresis. This process is widely used and has may applications. For example, it is used to analyze DNA molecules according to their resultant size after being digested by restriction enzymes. It is also used to analyze the products of a polymerase chain reaction (PCR).
Typically, electrophoresis separation is carried out in a separation matrix, such as a gel of agarose or acrylamide or a combination of the two. Usually, agarose gels are cast in open trays and form a slab whereas acrylamide gels are cast between two glass plates.
In order to effect the electrophoretic process, two opposite ends of the gels are exposed to a buffer solution, which is connected by electrodes, typically platinum, to an electrical power source. Once the electrical power source is switched on, the electric field forces negatively charged molecules to move towards the anode and positively charged molecules to move towards the cathode. The electrodes that are commonly used for electrophoresis are generally made of inert metals that induce water electrolysis in aqueous solutions, which produces hydroxyl ions at the cathode side and protons at the anode side. Therefore, large volumes of buffer are used in order to maintain the pH. In addition, due to the use of buffers of low salt concentration, large volumes of buffer are required to maintain the electric field.
DNA is negatively charged and therefore, in the agarose or acrylamide gels which provide sieving action, DNA molecules move towards the anode at a rate which depends on their size, wherein the smaller the molecules the faster they move.
In the electrophoretic separation of proteins, the proteins are often treated with an ionic detergent, such as sodium dodecylsulphate (SDS). The negatively charged dodecylsulphate anions interact with hydrophobic domains on the protein molecules, thus creating negatively charged protein/SDS complexes that undergo electrophoresis separation similar to DNA molecules.
Typically, it is desirable to visualize and to document the results of the electrophoretic separation test. In electrophoretic separation of DNA molecules, this has been done by immersing the gel slab after the electrophoretic separation has been completed in a solution of a fluorescent compound which emits visible light when exposed to a ultra violet (UV) light. A widely used compound is ethidium bromide.
Conventional electrophoretic systems are deficient in many respects, a few of which are listed below.
Prior art electrophoresis systems are a potential source of contamination to the working environment in which the tests are performed. The two major sources of contamination are ethidium bromide and PCR products. Ethidium bromide is a hazardous chemical due to its mutagenic activity. In addition, the environment is a potential source of contamination to the system, since PCR is an extremely sensitive method. In fact, a single molecule of DNA product from one PCR (out of the trillions of molecules being produced) may interfere with the subsequent PCR such that it will produce incorrect results.
Conventional electrophoresis is also deficient in other respects, one being that it is time consuming.
Many different gel separation materials have been disclosed, with different compositions, pH characteristics, voltage requirements, etc. The goal of most of the recent innovations in the field has been to provide an electrophoresis gel which can be used to perform a faster, more accurate, more stable, or more versatile electrophoresis separation.
U.S. Pat. No. 4,874,491 to Stalberg discloses an electophoresis system having a high concentration buffer containing gel.
U.S. Pat. No. 4,892,639 to Sarrine et al. discloses an electrophoresis plate with improved buffer circulation.
U.S. Pat. No. 5,045,164 to Tansamrit et al. discloses an electrophoresis plate having thickened ends as buffer reservoirs.
U.S. Pat. No. 5,209,831 to MacConnel discloses a bufferless disposable cassette having open ends and conductive film electrodes.
U.S. Pat. No. 5,407,552 to Lebacq and U.S. Pat. No. 5,411,657 to Leka disclose open electrophoresis devices requiring a buffer tank for operation.
U.S. Pat. No. 6,096,182 to Updyke et al. discloses an electrophoresis gel at a neutral pH. The advantage of producing such a gel is that the gel system is stable, with reduced reactivity and increased shelf life.